Phase state and surface topography of palmitoyl-ceramide monolayers

In cell biology (and in many biophysical) studies there is a natural tendency to consider ceramide as a highly condensed, solid-type lipid conferring rigidity and close packing to biomembranes. In the present work we advanced the understanding of the phase behavior of palmitoyl-ceramide restricted to a planar interface using Langmuir monolayers under strictly controlled and known surface packing conditions. Surface pressure-molecular area isotherms were complemented with molecular area-temperature isobars and with observations of the surface topography by Brewster Angle Microscopy. The results described herein indicate that palmitoyl-ceramide can exhibit expanded, as well as condensed phase states. Formation of three phases was found, depending on the surface pressure and temperature: a solid (1.80nm thick), a liquid-condensed (1.73nm thick, likely tilted) and a liquid-expanded (1.54nm thick) phase over the temperature range 5-62 °C. A large hysteretic behavior is observed for the S phase monolayer that may indicate high resistance to domain boundary deformation. A second (or higher) order S→LC phase transition is observed at about room temperature while a first order LC→LE transition occurs in a range of temperature encompassing the physiological one (observed above 30 °C at low surface pressure). This phase behavior broadens the view of ceramide as a type of lipid not-always-rigid but able to exhibit polymorphic properties.Fil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Maggio, Bruno. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin

Ordered-disordered domain coexistence in ternary lipid monolayers activates sphingomyelinase by clearing ceramide from the active phase

We explored the action of sphingomyelinase (SMase) on ternary monolayers containing phosphatidylcholine, sphingomyelin (SM) and dihydrocholesterol, which varied along a single tie line of phase coexistence. SMase activity exhibited a higher rate and extent of hydrolysis when the film is within the liquid-expanded (LE)/liquid-ordered (LO) coexistence range, compared to monolayers in the full LO phase. Since Alexa-SMase preferably adsorbs to the LE phase and there was no direct correlation found between enzymatic activity and domain borders, we postulate that the LE phase is the active phase for ceramide (Cer) generation. The enzymatically generated Cer was organized in different ways depending on the initial LE/LO ratio. The action of SMase in Chol-poor monolayers led to the formation of Cer-enriched domains, while in Chol-rich monolayers it resulted in the incorporation of Cer in the LO phase and the formation of new Chol- and Cer-enriched domains. The following novel mechanism is proposed to provide an explanation for the favored action of SMase on interfaces that exhibit an LE-LO phase coexistence: the LO phase sequesters the product Cer causing its depletion from the more enzyme-susceptible LE phase, thus decreasing inhibition by the reaction product. Furthermore, LO domains function as a substrate reservoir by allowing a rapid exchange of the substrate from this phase to the SM-depleted LE phase.Fil: Ale, Elisa Carmen. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Maggio, Bruno. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Química del Sur. Universidad Nacional del Sur. Departamento de Química. Instituto de Química del Sur; ArgentinaFil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin

Implementación de actividades de laboratorios en el nivel educativo primario

En este trabajo comentamos la experiencia adquirida durante el proyecto de articulación financiado por la Facultad de Ciencias Químicas de la Universidad Nacional de Córdoba durante el 2017 titulado "Implementación de actividades de laboratorios en Ciencias Naturales en el Nivel Educativo Primario". El proyecto se focalizó en la implementación de un laboratorio ambulante (que pueda ser transportado a las aulas) que abarque las distintas disciplinas de las Ciencias Naturales y en el diseño de Actividades didácticas en los niveles de 4to, 5to y 6to grado, en conjunto con docentes de diferentes escuelas, aprovechando el material de Laboratorio y equipamiento suministrado por la Presidencia de la Nación en el "Plan Nacional de enseñanza de las Ciencias Naturales para Escuela Primaria (año 2014), Modulo B". Se abordó la problemática a través de talleres con docentes de cuatro Escuelas Provinciales de la Provincia de Córdoba de nivel primario. Aquí describimos el desarrollo del proyecto, el porcentaje de su cumplimiento y los obstáculos atravesados. El resultado de la experiencia del trabajo conjunto se plasmó en el libro "Guía de actividades de laboratorios en ciencias naturales para escuelas primarias", escrito por las Dras. María Laura Fanani y Natalia Wilke, el cual tiene como objetivo servir de consulta para el desarrollo de las actividades de laboratorio en futuros ciclos lectivos.Fil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Wilke, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin

Miltefosine inhibits the membrane remodeling caused by phospholipase action by changing membrane physical properties

Miltefosine (hexadecylphosphocholine or HePC) is an alkylphosphocholine approved for the treatment of visceral and cutaneous Leishmaniasis. HePC exerts its effect by interacting with lipid membranes and affecting membrane-dependent processes. The molecular geometry of HePC suggests that the pharmacological function of HePC is to alter membrane curvature. As a model system, we studied the enzyme production in model membranes of diacylglycerol (DAG) or ceramide (CER), lipids involved in cell signaling which alter the structure of membranes. Here, we studied the effect of HePC on changes in phospholipase activity and on the effect that the lipid products have on the curvature and fusogenicity of membranes where they accumulate. Our results indicate that HePC inhibits the long-time restructuring of membranes, characteristic of the DAG and CER enzyme formation processes. In addition, the drug also reduces the fusogenicity of phospholipase-derived products. We postulate that the effect of HePC is due to a non-specific geometric compensation of HePC to the inverted cone-shape of DAG and CER products, acting as a relaxation agent of membrane curvature stress. These data are important for understanding the mechanism of action by which HePC regulates the lipid metabolism and signal transduction pathways in which these enzymes are involved.Fil: Zulueta Díaz, Yenisleidy de Las Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica; ArgentinaFil: Ambroggio, Ernesto Esteban. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin

Impact of sphingomyelin acyl chain (16:0 vs 24:1) on the interfacial properties of Langmuir monolayers: a PM-IRRAS study

Membrane structure is a key factor for the cell`s physiology, pathology, and therapy. Evaluating the importance of lipid species such as N-nervonoyl sphingomyelin (24:1-SM) —able to prevent phase separation— to membrane structuring remains a formidable challenge. This is the first report in which polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS) is applied to investigate the lipid-lipid interactions in 16:0 vs 24:1-SM monolayers and their mixtures with 1,2-Dioleoyl-sn-glycero-3-phosphocholine (DOPC) and cholesterol (Chol) (DOPC/SM/Chol 2:1:1). From the results we inferred that the cis double bond (Δ15) in 24:1-SM molecule diminishes intermolecular H-bonding and chain packing density compared to that of 16:0-SM. In ternary mixtures containing 16:0-SM, the relative intensity of the two components of the Amide I band reflected changes in the H-bonding network due to SM-Chol interactions. In contrast, the contribution of the main components of the Amide I band in DOPC/24:1-SM/Chol remained as in 24:1-SM monolayers, with a larger contribution of the non-H-bonded component. The most interesting feature in these ternary films is that the Cdouble bondO stretching mode of DOPC appeared with an intensity similar to that of SM Amide I band in DOPC/16:0-SM/Chol monolayers (a two-phase [Lo/Le] system), whereas an extremely low intensity of the Cdouble bondO band was detected in DOPC/24:1-SM/Chol monolayers (single Le phase). This is evidence that the unsaturation in 24:1-SM affected not only the conformational properties of acyl chains but also the orientation of the chemical groups at the air/water interface. The physical properties and overall H-bonding ability conferred by 24:1-SM may have implications in cell signaling and binding of biomolecules.Fil: Vázquez, Romina Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner". Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner"; Argentina. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Química; ArgentinaFil: Daza Millone, Maria Antonieta. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner". Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner"; ArgentinaFil: Pavinatto, Felippe J.. Universidade de Sao Paulo; BrasilFil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentina. Universidad Catolica de Córdoba. Facultad de Medicina. Departamento de Química Biologica; ArgentinaFil: Oliveira, Osvaldo N. Jr.. Universidade de Sao Paulo; BrasilFil: Vela, Maria Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; ArgentinaFil: Maté, Sabina María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner". Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Instituto de Investigaciones Bioquímicas de La Plata "Prof. Dr. Rodolfo R. Brenner"; Argentin

Atypical surface behavior of ceramides with nonhydroxy and 2-hydroxy very long-chain (C28-C32) PUFAs

Unique species of ceramide (Cer) with very-long-chain polyunsaturated fatty acid (VLCPUFA), mainly 28?32 carbon atoms, 4?5 double bonds, in nonhydroxy and 2-hydroxy forms (n-V Cer and h-V Cer, respectively), are generated in rat spermatozoa from the corresponding sphingomyelins during the acrosomal reaction. The aim of this study was to determine the properties of these sperm-distinctive ceramides in Langmuir monolayers. Individual Cer species were isolated by HPLC and subjected to analysis of surface pressure, surface potential, and Brewster angle microscopy (BAM) as a function of molecular packing. In comparison with known species of Cer, n-V Cer and h-V Cer species showed much larger mean molecular areas and increased molecular dipole moments in liquid expanded phases, which suggest bending and partial hydration of the double bonded portion of the VLCPUFA. The presence of the 2-hydoxyl group induced a closer molecular packing in h-V Cer than in their chain-matched n-V Cer. In addition, all these Cer species showed liquid-expanded to liquid-condensed transitions at room temperature. Existence of domain segregation was confirmed by BAM. Additionally, thermodynamic analysis suggests a phase transition close to the physiological temperature for VLCPUFA-Cers if organized as bulk dispersions.Fil: Peñalva, Daniel Alejandro. Consejo Nacional de Investigaciones Cientificas y Técnicas. Centro Científico Tecnológico Bahia Blanca. Instituto de Investigaciones Bioquímicas Bahia Blanca (i); ArgentinaFil: Oresti, Gerardo Martin. Consejo Nacional de Investigaciones Cientificas y Técnicas. Centro Científico Tecnológico Bahia Blanca. Instituto de Investigaciones Bioquímicas Bahia Blanca (i); ArgentinaFil: Dupuy, Fernando Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones En Química Biológica de Córdoba (p); ArgentinaFil: Antollini, Silvia Susana. Consejo Nacional de Investigaciones Cientificas y Técnicas. Centro Científico Tecnológico Bahia Blanca. Instituto de Investigaciones Bioquímicas Bahia Blanca (i); ArgentinaFil: Maggio, Bruno. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones En Química Biológica de Córdoba (p); ArgentinaFil: Aveldaño, Marta Isabel. Consejo Nacional de Investigaciones Cientificas y Técnicas. Centro Científico Tecnológico Bahia Blanca. Instituto de Investigaciones Bioquímicas Bahia Blanca (i); ArgentinaFil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones En Química Biológica de Córdoba (p); Argentin

Self-assembled nanostructures of L-ascorbic acid alkyl esters support monomeric amphotericin B

Amphotericin B (AmB) is a highly effective antimicrobial, with broad antimycotic and antiparasitic effect. However, AmB poor water-solubilisation and aggregation tendency limits its use for topical applications. We studied the capacity of nanostructures formed by alkyl esters of L-ascorbic acid (ASCn) to solubilise AmB and tested the relationship between the prevalence of the monomeric form of AmB and its effectiveness as antimicrobial agent.Fil: Nocelli, Natalia Estefanía. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Biología Molecular; ArgentinaFil: Zulueta Díaz, Yenisleidy de Las Mercedes. Universidad Nacional de Córdoba; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Millot, Marine. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica; ArgentinaFil: Colazo, María Luz. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Química Biológica; ArgentinaFil: Vico, Raquel Viviana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones en Físico-química de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Instituto de Investigaciones en Físico-química de Córdoba; ArgentinaFil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin

Impact of sphingomyelin acyl chain (16:0 vs 24:1) on the interfacial properties of Langmuir monolayers: A PM-IRRAS study

Membrane structure is a key factor for the cell`s physiology, pathology, and therapy. Evaluating the importance of lipid species such as N-nervonoyl sphingomyelin (24:1-SM) -able to prevent phase separation- to membrane structuring remains a formidable challenge. This is the first report in which polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS) is applied to investigate the lipid-lipid interactions in 16:0 vs 24:1-SM monolayers and their mixtures with 1,2-Dioleoyl-sn-glycero-3-phosphocholine (DOPC) and cholesterol (Chol) (DOPC/SM/Chol 2:1:1). From the results we inferred that the cis double bond (Δ15) in 24:1-SM molecule diminishes intermolecular H-bonding and chain packing density compared to that of 16:0-SM. In ternary mixtures containing 16:0-SM, the relative intensity of the two components of the Amide I band reflected changes in the H-bonding network due to SM-Chol interactions. In contrast, the contribution of the main components of the Amide I band in DOPC/24:1-SM/Chol remained as in 24:1-SM monolayers, with a larger contribution of the non-H-bonded component. The most interesting feature in these ternary films is that the CO stretching mode of DOPC appeared with an intensity similar to that of SM Amide I band in DOPC/16:0-SM/Chol monolayers (a two-phase [Lo/Le] system), whereas an extremely low intensity of the CO band was detected in DOPC/24:1-SM/Chol monolayers (single Le phase). This is evidence that the unsaturation in 24:1-SM affected not only the conformational properties of acyl chains but also the orientation of the chemical groups at the air/water interface. The physical properties and overall H-bonding ability conferred by 24:1-SM may have implications in cell signaling and binding of biomolecules.Instituto de Investigaciones Bioquímicas de La PlataInstituto de Investigaciones Fisicoquímicas Teóricas y Aplicada

The many faces (and phases) of ceramide and sphingomyelin II – binary mixtures

A rather widespread idea on the functional importance of sphingolipids in cell membranes refers to the occurrence of ordered domains enriched in sphingomyelin and ceramide that are largely assumed to exist irrespective of the type of N-acyl chain in the sphingolipid. Ceramides and sphingomyelins are the simplest kind of two-chained sphingolipids and show a variety of species, depending on the fatty acyl chain length, hydroxylation, and unsaturation. Abundant evidences have shown that variations of the N-acyl chain length in ceramides and sphingomyelins markedly affect their phase state, interfacial elasticity, surface topography, electrostatics, and miscibility, and that even the usually conceived “condensed” sphingolipids and many of their mixtures may exhibit liquid-like expanded states. Their lateral miscibility properties are subtlety regulated by those chemical differences. Even between ceramides with different acyl chain length, their partial miscibility is responsible for a rich two-dimensional structural variety that impacts on the membrane properties at the mesoscale level. In this review, we will discuss the miscibility properties of ceramide, sphingomyelin, and glycosphingolipids that differ in their N-acyl or oligosaccharide chains. This work is a second part that accompanies a previous overview of the properties of membranes formed by pure ceramides or sphingomyelins, which is also included in this Special Issue.Fil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Maggio, Bruno. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin

Regulation of phase boundaries and phase-segregated patterns in model membranes

Demixing of components has long been described in model membranes. It is a consequence of non-ideal lateral interactions between membrane components, and it causes the presence of segregated phases, forming patches (domains) of different properties, thus introducing heterogeneity into the membrane. In the present review we first describe the processes through which domains are generated, how they grow, and why they are rounded, striped or fractal-like, as well as why they get distributed forming defined patterns. Next, we focus on the effect of an additive on a lipid mixture, which usually induces shifts in demixing points, thus stabilizing or destabilizing the phase-segregated state. Results found for different model membranes are summarized, detailing the ways in which phase segregation and the generated patterns may be modulated. We focus on which are, from our viewpoint, the most relevant regulating factors affecting the surface texture observed in model membranes. This article is part of a Special Issue entitled: Emergence of Complex Behavior in Biomembranes edited by Marjorie Longo.Fil: Fanani, Maria Laura. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; ArgentinaFil: Wilke, Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Centro de Investigaciones en Química Biológica de Córdoba. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Centro de Investigaciones en Química Biológica de Córdoba; Argentin